The filamentous fungus Aspergillus niger is widely exploited by the fermentation industry for the production of enzymes and organic acids, particularly citric acid. We sequenced the 33.9-megabase genome of A. niger CBS 513.88, the ancestor of currently used enzyme production strains. A high level of synteny was observed with other aspergilli sequenced. Strong function predictions were made for 6,506 of the 14,165 open reading frames identified. A detailed description of the components of the protein secretion pathway was made and striking differences in the hydrolytic enzyme spectra of aspergilli were observed. A reconstructed metabolic network comprising 1,069 unique reactions illustrates the versatile metabolism of A. niger. Noteworthy is the large number of major facilitator superfamily transporters and fungal zinc binuclear cluster transcription factors, and the presence of putative gene clusters for fumonisin and ochratoxin A synthesis.

Starch is a major storage product of many economically important crops such as wheat, rice, maize, tapioca, and potato. A large-scale starch processing industry has emerged in the last century. In the past decades, we have seen a shift from the acid hydrolysis of starch to the use of starch-converting enzymes in the production of maltodextrin, modified starches, or glucose and fructose syrups. Currently, these enzymes comprise about 30% of the world's enzyme production. Besides the use in starch hydrolysis, starch-converting enzymes are also used in a number of other industrial applications, such as laundry and porcelain detergents or as anti-staling agents in baking. A number of these starch-converting enzymes belong to a single family: the alpha-amylase family or family13 glycosyl hydrolases. This group of enzymes share a number of common characteristics such as a (beta/alpha)(8) barrel structure, the hydrolysis or formation of glycosidic bonds in the alpha conformation, and a number of conserved amino acid residues in the active site. As many as 21 different reaction and product specificities are found in this family. Currently, 25 three-dimensional (3D) structures of a few members of the alpha-amylase family have been determined using protein crystallization and X-ray crystallography. These data in combination with site-directed mutagenesis studies have helped to better understand the interactions between the substrate or product molecule and the different amino acids found in and around the active site. This review illustrates the reaction and product diversity found within the alpha-amylase family, the mechanistic principles deduced from structure-function relationship structures, and the use of the enzymes of this family in industrial applications.

Although abundant geochemical data indicate that anaerobic methane oxidation occurs in marine sediments, the linkage to specific microorganisms remains unclear. In order to examine processes of methane consumption and oxidation, sediment samples from mud volcanoes at two distinct sites on the Mediterranean Ridge were collected via the submersible Nautile. Geochemical data strongly indicate that methane is oxidized under anaerobic conditions, and compound-specific carbon isotope analyses indicate that this reaction is facilitated by a consortium of archaea and bacteria. Specifically, these methane-rich sediments contain high abundances of methanogen-specific biomarkers that are significantly depleted in 13 C (␦ 13 C values are as low as ؊95‰). Biomarkers inferred to derive from sulfate-reducing bacteria and other heterotrophic bacteria are similarly depleted. Consistent with previous work, such depletion can be explained by consumption of 13 Cdepleted methane by methanogens operating in reverse and as part a consortium of organisms in which sulfate serves as the terminal electron acceptor. Moreover, our results indicate that this process is widespread in Mediterranean mud volcanoes and in some localized settings is the predominant microbiological process.Methane can have a stronger greenhouse effect than CO 2 , and recent work has highlighted its potential climatic impact on glacial timescales (24) and in relation to major geologic events (8,12). Consequently, controls on methane production and consumption are important concerns in the evaluation of past and future climate change. In marine sediments, anaerobic methane oxidation could be the dominant pathway for methane consumption (2,4,6,15,25,26), but the organisms involved have not been isolated and the mechanism remains controversial. Methanogenic archaea operating in reverse (9,11,14) or novel, previously uncharacterized archaea (13) have been proposed to play a vital role, but current evidence remains ambiguous.Methane tends to be highly depleted in 13 C, and organisms that consume methane either directly or indirectly via heterotrophic consumption of methanotroph biomass will be similarly depleted in 13 C. For this study, we determined distributions and carbon isotope abundances of organic components in methane-rich mud volcano sediments of the Eastern Mediterranean Ridge. In particular, we examined the ␦ 13 C values of compounds derived from specific organisms (i.e., biomarkers) presumed to play a major role in methane oxidation, including methanogens, aerobic methanotrophs, sulfate-reducing bacteria, and anaerobic heterotrophic bacteria. From these results, we evaluate the controls on and microorganisms responsible for methane oxidation in these sediments and the conditions under which methane oxidation occurs. MATERIALS AND METHODSSamples. Using a bordeaux core, samples were collected from surface sediments (upper 30 cm) of several mud volcano flows (mud breccias) in the Olimpi (Milano and Napoli mud volcanoes) and Anaximander (Amsterdam mud volcano) fi...

Members of the genera Streptococcus and Leuconostoc synthesize various a-glucans (dextran, alternan and mutan). In Lactobacillus, until now, the only glucosyltransferase (GTF) enzyme that has been characterized is gtfA of Lactobacillus reuteri 121, the first GTF enzyme synthesizing a glucan (reuteran) that contains mainly a-(1R4) linkages together with a-(1R6) and a-(1R4,6) linkages. Recently, partial sequences of glucansucrase genes were detected in other members of the genus Lactobacillus. This paper reports, for the first time, isolation and characterization of dextransucrase and mutansucrase genes and enzymes from various Lactobacillus species and the characterization of the glucan products synthesized, which mainly have a-(1R6)-and a-(1R3)-glucosidic linkages. The four GTF enzymes characterized from three different Lb. reuteri strains are highly similar at the amino acid level, and consequently their protein structures are very alike. Interestingly, these four Lb. reuteri GTFs have relatively large N-terminal variable regions, containing RDV repeats, and relatively short putative glucan-binding domains with conserved and less-conserved YG-repeating units. The three other GTF enzymes, isolated from Lactobacillus sakei, Lactobacillus fermentum and Lactobacillus parabuchneri, contain smaller variable regions and larger putative glucan-binding domains compared to the Lb. reuteri GTF enzymes.